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- EMDB-4017: A 4.2 Angstrom structure of HIV-1 CA-SP1 from immature HIV-1 particles -

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Basic information

Entry
Database: EMDB / ID: EMD-4017
TitleA 4.2 Angstrom structure of HIV-1 CA-SP1 from immature HIV-1 particles
Map data
Sample
  • Virus: Human immunodeficiency virus 1
    • Protein or peptide: HIV-1 capsid-SP1 from immature HIV-1 particles
Biological speciesHuman immunodeficiency virus 1
Methodsubtomogram averaging / cryo EM / Resolution: 4.2 Å
AuthorsSchur FKM / Obr M / Hagen WJH / Wan W / Arjen JJ / Kirkpatrick JM / Sachse C / Kraeusslich H-G / Briggs JAG
CitationJournal: Science / Year: 2016
Title: An atomic model of HIV-1 capsid-SP1 reveals structures regulating assembly and maturation.
Authors: Florian K M Schur / Martin Obr / Wim J H Hagen / William Wan / Arjen J Jakobi / Joanna M Kirkpatrick / Carsten Sachse / Hans-Georg Kräusslich / John A G Briggs /
Abstract: Immature HIV-1 assembles at and buds from the plasma membrane before proteolytic cleavage of the viral Gag polyprotein induces structural maturation. Maturation can be blocked by maturation ...Immature HIV-1 assembles at and buds from the plasma membrane before proteolytic cleavage of the viral Gag polyprotein induces structural maturation. Maturation can be blocked by maturation inhibitors (MIs), thereby abolishing infectivity. The CA (capsid) and SP1 (spacer peptide 1) region of Gag is the key regulator of assembly and maturation and is the target of MIs. We applied optimized cryo-electron tomography and subtomogram averaging to resolve this region within assembled immature HIV-1 particles at 3.9 angstrom resolution and built an atomic model. The structure reveals a network of intra- and intermolecular interactions mediating immature HIV-1 assembly. The proteolytic cleavage site between CA and SP1 is inaccessible to protease. We suggest that MIs prevent CA-SP1 cleavage by stabilizing the structure, and MI resistance develops by destabilizing CA-SP1.
History
DepositionJun 9, 2016-
Header (metadata) releaseJul 13, 2016-
Map releaseJul 20, 2016-
UpdateNov 28, 2018-
Current statusNov 28, 2018Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.115
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.115
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_4017.png

Downloads & links

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Map

FileDownload / File: emd_4017.map.gz / Format: CCP4 / Size: 27 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.35 Å/pix.
x 192 pix.
= 259.2 Å		1.35 Å/pix.
x 192 pix.
= 259.2 Å		1.35 Å/pix.
x 192 pix.
= 259.2 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.35 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.115 / Movie #1: 0.115
Minimum - Maximum-0.26954892 - 0.41738236
Average (Standard dev.)0.0014261577 (±0.044672597)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions192192192
Spacing192192192
CellA=B=C: 259.2 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.351.351.35
M x/y/z192192192
origin x/y/z0.0000.0000.000
length x/y/z259.200259.200259.200
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS192192192
D min/max/mean-0.2700.4170.001

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Supplemental data

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Sample components

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Entire : Human immunodeficiency virus 1

EntireName: Human immunodeficiency virus 1
Components
  • Virus: Human immunodeficiency virus 1
    • Protein or peptide: HIV-1 capsid-SP1 from immature HIV-1 particles

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Supramolecule #1: Human immunodeficiency virus 1

SupramoleculeName: Human immunodeficiency virus 1 / type: virus / ID: 1 / Parent: 0 / Macromolecule list: all
Details: Immature protease defective (D25A) viruses were purified from HEK293T cells
NCBI-ID: 11676 / Sci species name: Human immunodeficiency virus 1 / Virus type: VIRION / Virus isolate: OTHER / Virus enveloped: Yes / Virus empty: No
Host (natural)Organism: Homo sapiens (human)

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Macromolecule #1: HIV-1 capsid-SP1 from immature HIV-1 particles

MacromoleculeName: HIV-1 capsid-SP1 from immature HIV-1 particles / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
SequenceString: SPRTLNAWVK VVEEKAFSPE VIPMFSALSE GATPQDLNTM LNTVGGHQAA MQMLKETINE EAAEWDRLH PVHAGPIAPG QMREPRGSDI AGTTSTLQEQ IGWMTHNPPI PVGEIYKRWI I LGLNKIVR MYSPTSILDI RQGPKEPFRD YVDRFYKTLR AEQASQEVKN ...String:
SPRTLNAWVK VVEEKAFSPE VIPMFSALSE GATPQDLNTM LNTVGGHQAA MQMLKETINE EAAEWDRLH PVHAGPIAPG QMREPRGSDI AGTTSTLQEQ IGWMTHNPPI PVGEIYKRWI I LGLNKIVR MYSPTSILDI RQGPKEPFRD YVDRFYKTLR AEQASQEVKN WMTETLLVQN AN PDCKTIL KALGPGATLE EMMTACQGVG GPGHKARVLA EAMSQVT

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Experimental details

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Structure determination

Methodcryo EM
Processingsubtomogram averaging
Aggregation stateparticle

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Sample preparation

BufferpH: 7 / Component:
NameConcentration
PBS
Paraformaldehyde1.2 %

Details: Concentrated virus sample was resuspended in PBS and fixed with 1.2 % PFA for one hour on ice.
GridModel: C-flat / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Details: at 20 mA
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 15 K / Instrument: FEI VITROBOT MARK II
Details: 10nM colloidal gold was added to the sample prior to plunge freezing..

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: GIF Quantum LS / Energy filter - Lower energy threshold: 0 eV / Energy filter - Upper energy threshold: 20 eV
DetailsNanoprobe
Image recordingFilm or detector model: GATAN K2 QUANTUM (4k x 4k) / Detector mode: SUPER-RESOLUTION / Digitization - Dimensions - Width: 3710 pixel / Digitization - Dimensions - Height: 3838 pixel / Digitization - Frames/image: 10-12 / Number grids imaged: 1 / Average exposure time: 2.5 sec. / Average electron dose: 3.0 e/Å2
Details: Number of frames ranged from 10-12. Exposure time per tilt ranged from 2 to 3 seconds
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 5.0 µm / Nominal defocus min: 1.5 µm / Nominal magnification: 105000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

DetailsFrames were aligned using MotionCorr. Tilts in a tilt series were exposure filtered for cumulative electron dose. Tomograms were reconstructed using IMOD.
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C6 (6 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 4.2 Å / Resolution method: FSC 0.143 CUT-OFF / Software: (Name: AV3, TOM Toolbox) / Number subtomograms used: 100537
ExtractionNumber tomograms: 74 / Number images used: 747729
Details: Subtomograms were extracted from the surface of each particle according to the determined radius of the particle.
CTF correctionSoftware:
Namedetails
CTFFIND (ver. 4)CTF determination
CTFPHASEFLIPCTF correction within IMOD

Details: CTF correction was performed using the ctfphaseflip program in IMOD prior to backprojection.
Final angle assignmentType: OTHER / Software: (Name: AV3, TOM Toolbox) / Details: Cross-correlation based template matching

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